Inventions of an observation apparatus which generates a phase image on the basis of an interference image of an observation object flowing in a flow path with a fluid are disclosed in Patent Documents 1 to 3. For example, the flow path is a flow cell, the fluid is blood, and the observation object is a red blood cell, a white blood cell, a circulating tumor cell (hereinafter, referred to as “CTC”) in the blood, and the like. The CTC is a cell which is separated from a primary tumor tissue or a metastatic tumor tissue and enters the blood, and is found infinitesimally in peripheral blood of a solid cancer patient, and is involved in metastasis.
In recent years, the CTC has been actively studied. In clinical application fields, it has been reported that there is a certain relation between the number of CTCs per unit blood volume and a survival rate of a patient after certain years. Identifying and inspecting the circulating tumor cell are very useful to prognostic prediction of cancer, determination on chemotherapeutic efficacy, screening of cancer, gene therapy, and drug development support for anticancer agent or the like, and is very valuable to the clinical applications.
An object can generally be identified on the basis of size, shape, or color. However, the white blood cell and the CTC are not so different in size, and both are colorless and transparent, and therefore, both cells are hard to be identified from an image obtained by a bright field microscope. Further, a phase contrast microscope and a differential interference microscope are used to visualize a colorless and transparent cell, but there is a defect in quantitativity with respect to an optical thickness. Further, the focal depth of these microscopes is equal to or less than a thickness of the cell depending on a using objective lens, and therefore, only two-dimensional information can be obtained even though the cell is structured three-dimensionally, and thus the object is not possible to be identified.
In the observation apparatuses of the inventions disclosed in Patent Documents 1 to 3, the phase image is generated on the basis of the interference image of the observation object which flows in the flow path with the fluid, and a shape (an external form, a shape of a nucleus, or the like) of the observation object is analyzed on the basis of the phase image, and therefore, the CTC in the blood can be identified. The conventional observation apparatus splits light output from a light source into first split light and second split light, reflects or transmits the first split light by the observation object in the flow path, causes interference between the first split light and the second split light to repeatedly acquire two-dimensional interference images, and generates the phase image of the observation object on the basis of these two-dimensional interference images. In order to acquire the two-dimensional interference image, the conventional observation apparatus uses an imaging unit having a plurality of pixels two-dimensionally arranged on an imaging plane.